A novel positive feedback loop involving FASN/p-ERK1/2/5-LOX/LTB4/FASN sustains high growth of breast cancer cells.

Aim: To investigate the endogenous signaling pathways associated with high proliferation potential of breast cancer cells.

Methods: Breast cancer cell lines LM-MCF-7 and MCF-7 with high and low proliferation capability were used. The promoter activity of fatty acid synthase (FASN) was examined using luciferase reporter gene assay. The expression level of FASN mRNA was measured using RT-PCR and real time PCR, respectively. The level of leukotriene B4 (LTB4) was determined with ELISA. The expression levels of 5-lipoxygenase (5-LOX) was analyzed using RT-PCR and Western blot, respectively. 5-Bromo-20-deoxyuridine (BrdU) incorporation assay was used to study the proliferation of LM-MCF-7 and MCF-7 cells.

Results: The promoter activity of FASN was significantly higher in LM-MCF-7 cells than MCF-7 cells. Treatment of LM-MCF-7 cells with ERK1/2 inhibitor PD98059 (30-50 μmol/L) or LOX inhibitor NDGA (25 μmol/L) abolished the activation of FASN. Moreover, treatment of LM-MCF-7 cells with the specific 5-LOX inhibitor MK-886 (20-40 μmol/L) or 5-LOX siRNA (50-100 nmol/L) decreased the promoter activity of FASN. The level of LTB4, the final metabolite produced by 5-LOX, was significantly higher in LM-MCF-7 cells than MCF-7 cells. Administration of exogenous LTB4 (1-10 nmol/L) was able to stimulate the promoter activity of FASN in MCF-7 cells. Treatment of LM-MCF-7 cells with the FASN inhibitor cerulenin (10 μmol/L) reduced all the levels of p-ERK1/2, 5-LOX, and LTB4. Treatment of LM-MCF-7 cells with cerulenin, PD98059, or MK-886 abolished the proliferation. Administration of exogenous LTB4 (10 nmol/L) significantly increased BrdU incorporation in MCF-7 cells.

Conclusion: THESE results suggest a novel positive feedback loop involving FASN/p-ERK1/2/5-LOX/LTB4/FASN contributes to the sustaining growth of breast cancer LM-MCF-7 cells.